Method for the treatment of diseases due to defects the immune system

ABSTRACT

New crystalline organic salts of N,N&#39;-diacetyl cystine with immunomodulating effect, processes for their preparation, pharmaceutical compositions containing them and methods of their pharmacological use.

This application is a divisional of application Ser. No. 08/335,941,filed on Nov. 8. 1994, now U.S. Pat. No. 5,693,858, which is acontinuation of application Ser. No. 07/981,375, filed Nov. 25, 1992,issued as U.S. Pat. No. 5.383,904 on Jan. 31, 1995.

FIELD OF THE INVENTION

The present invention relates to organic salts of N,N'-diacetyl cystine,in the following referred to as DiNAc, with immunomodulating activity aswell as pharmaceutical compositions based on these salts and methods fortheir pharmacological use. The invention specifically concerns methodsto obtain crystalline, non-hygroscopic and chemically stable saltscontaining non-toxic organic cations, which are useful for therapy ofdiseases where a defect in the immune system is indicated.

BACKGROUND ART

N-Acetyl-L-cysteine is a well known compound which is routinely used asa therapeutic agent against chronic obstructive diseases and chronicbronchitis. Although the first patent was filed in 1964 (GB 954268) themechanism of action of the compound has not been established. It hasrecently been discovered that the corresponding disulfide ofN-Acetyl-L-cysteine, i.e. L-DiNAc, acts as a potent immunostmulator(Swedish patent application no SE 9002067-8), showing an activitycomparable to contemporary immunostimulants such as sodium diethyldithiocarbamate or 2,2'-dithiobisethanol.

Routes for the preparation of DiNAc have been reported in severalpatents (U.S. Pat. Nos. 4,827,016, 4,724,239 and 4,708,965, EuropeanPatent No 300100 and German Patent No 2326444). DiNAc is, however,amorphous and hygroscopic, so that it is difficult to isolate andformulate into pharmaceutical compositions and its administration isnormally only in the form of aqueous solutions. Most salts of DiNAc withinorganic or organic cations share the same unfavourable physicalproperties with the free diacid. Examples of salts of other,sulfur-containing, amino acids have appeared in the patent literature(U.S. Pat. No. 3,647,834, JP patent No 56155298 and FR patent No8106592)

DISCLOSURE OF THE INVENTION

We have now surprisingly found that a few salts of DiNAc with organicbases exhibit a favourable combination of non-hygroscopicity andcrystallinity which permits the isolation and formulation of these saltsin solid form and allows them to be administered by inhalation in solidform, or in other dry formulations, should this be clinically desirable.

The present invention provides organic salts of DiNAc, having theformula la or lb: ##STR1## where R⁺ and R²⁺ is a mono- or diprotonatedorganic amine respectively. The organic amine is selected from lysinium,ethylenediaminium, N,N'-dibenzylethylenediaminium,N-benzyl-2-phenylethylaminium, piperazinium and 1 -adamantanaminium.

Lysinium can be in its D- or L-form. Most preferred is the L-form.

The invention includes hydrated and solvated salts, e.g. solvated withlower alkanols. The invention includes salts of DiNAc in its individualisomers, i.e. D-, L- and meso-forms as well as in its racemic form. Mostpreferred are the L-forms of these salts.

We have found that the new salts of the invention fulfil therequirements of ease of crystallization, non-hygroscopicity and chemicalstability while still retaining the immunomodulating activity of DiNAc,and are thus medically useful.

This invention thus provides compounds with advantageous properties forthe treatment of diseases where an anergy of the immune response or anaberrant immune response or an ineffective host defence can besuspected. Such diseases include chronic bronchitis, where a reductionof the rate of exacerbations has previously been reported with immuneresponse modifiers such as Biostim (Radermecker, M. et al. Int. J.Immunopharmac. 10, 913-917, 1988; Scheffer, J. et al. Arzneim.Forsch/Drug Res. 41, 815820, 1991), Ribomunyl and BronchoVaxom (Paupe,J. Respiration 58, 150-154, 1991) as well as with N-Acetylcysteine (SeeBergstrand, H. et al J. Free Radic. Biol. Med. 2, 119-127, 1986).

Such diseases also include certain forms of malignant diseases. Thus,numerous research institutes round the world aim at finding ways ofstimulating the immune response of patients with various forms ofmalignant diseases and numerous reviews in the literature deal with thisapproach (Stevenson, F. K. FASEB J 5:2250-2257, 1991). To mention oneexample patients with intracranial tumours (gliomas) exhibit a profounddecrease in immunity possibly due to a defect in the secretion of IL-2as well as the expression of IL-2 receptors in T cells from suchpatients (Roszman, T. et al. Immunology Today 12, 370 374, 1991).Moreover, a significant adjuvant effect in immunotherapy of melanoma andcolon carcinoma has been documented for the immunostimulator Levamisole(Van Wauwe, J. and Janssen, P. A. J. Int J. Immunopharmac. 13, 3-9,1991) and immunotherapy with IL-2 in vivo or treatment of patientslymphokine activated killer cells with IL-2 ex vivo has caused theregression of cancer in selected patients (Rosenberg, S. A. ImmunologyToday 9, 58-, 1988). The malignant diseases at which the compounds I canbe expected to have advantageous effects include tumours of mesenchymalorigin such as sarcomas like fibrosarcoma, myxosarcoma, liposarcoma,chondrosarcoma, osteogenic sarcoma or chordosarcoma, sarcomas likeangiosarcoma, endotheliosarcoma, lymphangiosarcoma, synoviosarcoma ormesotheliosarcoma, leukemias and lymphomas like granulocytic leukemia,monocytic leukemia, lymphocytic leukemia, malignant lymphoma,plasmocytoma, reticulum cell sarcoma or Hodkins disease, sarcomas likeleiomyosarcoma or rhabdomysarcoma, tumours of epithelial origin(Carcinomas) like squamous cell carcinoma, basal cell carcinoma, sweatgland carcinoma, sebaceous gland carcinoma, adenocarcinoma, papillarycarcinoma, papillary adenocarcinoma, cystadenocarcinoma, medullarycarcinoma, undifferentiated carcinoma, bronchogenic carcinoma, melanoma,renal cell carcinoma, hepatoma-liver cell carcinoma, bile ductcarcinoma-cholangiocarcinoma, transitional cell carcinoma,choriocarcinoma, semonoma or embryonal carcinoma, tumours of the centralnervous system like glioma, meningoma, medulloblastoma, schwannoma orependymoma.

Moreover, the compounds also have advantageous properties for treatmentof chronic infections such as herpes, aphtous stomatitis and minimalchange syndrome where clinical improvement has previously been reportedby treatment with an immunostimulator such as Levamisole as well asother chronic inflammatory diseases in the urinary tract or in ear, noseor throat, which benefit from treatment with immunostimulators such asBiostim, Broncho-Vaxom and Ribomunyl, or an HIV infection or AIDS.

Moreover, an impairment, a defect or an imbalance of the immune responsehas also been postulated to exist at atopic diseases such as atopicdermatitis, rhinitis and asthma (Katz, D. H. Transplantation Reviews 41,77-108, 1977). Since theoretical considerations suggest that stimulationof an immune response would possibly be the best way of restoringimbalances and autoimmunity (Varela, F. J. and Coutinho, A. ImmunologyToday 12, 159-166, 1991), the compounds can also be expected to haveadvantageous properties for treatment of asthma, rhinitis, atopicdermatitis and autoimmune diseases like non-obese diabetes, systemiclupus erythematosus, sclerodermia, Sjogren's syndrome, dermatomyositisor multiple sclerosis, rheumatoid arthritis and possibly psoriasis.

Moreover, the compounds, due to their immune stimulating properties, canbe expected to have advantageous properties as adjuvants in variousforms of vaccine preparations.

Finally, the compounds can be expected to have advantageous propertiesfor treatment of atherosclerosis whether or not it will influence aputative inflammatory process in this condition (Hansson. G. K. et al.Proc. Nat. Acad. Sci. USA 88,10530, 1991).

The importance of immunostimulatory drugs on tumour outgrowth isillustrated in our relevant tumour outgrowth test systems. Theexperimental rat tumour models described below reflect very well thetumourostatic potency of the compounds of this invention in comparisonwith the well established immunostimulatory drug Levamisole, and theyreflect clinically very well a fastly growing tumour, the mammarycarcinoma, and a slowly progressively growing tumour, the glioma, and inboth systems the drug has an excellent static effect on tumouroutgrowth.

Particularly suitable for treatment with the compounds of this inventionare:

Malignancies such as melanoma, mammary carcinoma, gastrointestinalcarcinoma, glioma, bladder carcinoma and squamous cell carcinoma of theneck and head region;

Infections such as chronic bronchitis, hepatitis, post-infectious anergyand aquired immune deficiencies such as AIDS;

Posttraumatic immunological arergy, and

Purported autoimmune diseases such as rheumatoid arthritis, multiplesclerosis and psoriasis.

Effective doses for treatment of the diseases above are in the range of0.1-100 mg, preferably 1.0-60 mg daily.

Methods of Preparation

The organic salts of formula la and lb are generally prepared by mixingDiNAc and the organic base, as defined above, each dissolved ordispersed in a solvent or solvent mixture. Solvents such as water,alcohols, glycols, ketones, amides, sulphoxides or other polar solventsor solvent mixtures may be used. The salt either precipitates directlyfrom the reaction mixture, or is obtained by the addition of a lesspolar solvent or by evaporation or lyophilisation. The reaction isperformed at elevated temperature or room temperature, depending on thesolubility in the medium. Alternatively, the salt can be prepared byoxidation of the appropriate N-acetyl cysteine salt in aqueous oralcoholic solution, followed by precipitation as above. The oxidationmay be effected either chemically, using e.g. hydrogen peroxide orhalogen, or electrochemically.

Pharmaceutical Formulations

According to the present invention the compounds of the formula la andlb can be formulated for administration by inhalation as well as byother routes, e.g. orally or topically with or without apharmaceutically acceptable carrier.

The substance can be inhaled from a pressurized metered dose inhaler,from a dry powder inhaler, e.g. Turbuhaler® or from a dry powder inhalerutilizing gelatine, plastic or other capsules. Non-toxic and chemicallyinert substances e.g. lactose, trehalose, mannitol or glucose, can beadded to the powdered substance.

The substance can also be administered orally in the form of capsules ortablets where the capsule, the tablet or the active substance itself canbe coated or non-coated. This coating may consist of for examplehydroxypropyl cellulose and aerosil in isopropanol or other suitablesolvents. A non-toxic and chemically inert substance can be added to thepowdered substance in order to obtain desirable physical orpharmaceutical properties.

Parenteral administration of the new compounds is also possible.

Pharmacological Experiments

The ability of the compound di-L-lysinium-N,N'-diacetyl-L-cystinate (la;R=lysine) to modulate immune responses is illustrated by its efficacy inthe animal delayed type hypersensitivity (DTH) test in the mouse.

Both male and female Balb/c mice, obtained from Bomholtsgaard (Denmark),were used at the weight of 18-20 gram.4-Ethoxymethylene-2-phenyloxazolin-5-one (OXA) was purchased from BDH(England) and served as the antigen in this test.

The mice were sensitized, Day 0, by epicutaneous application of 150 μlof an absolute ethanol-acetone (3:1) solution containing 3% OXA on theshaved abdomen. Treatment with the disulphide salt or vehicle (0.9%NaCl) was initiated by oral feeding immediately after sensitization andcontinued once daily until Day 6. Seven days (Day 6) after thesensitization, both ears of all mice were challenged on both sides bytopical application of 20 μl 1 % OXA dissolved in peanut oil. Earthickness was measured prior to and 24 or 48 hours after challenge usingan Oditest spring calliper. Challenges and measurements were performedunder light pentobarbital anaesthesia.

The intensity of the DTH reactions was expressed according to theformula: T_(t24/48) -T_(t0) μm units, where t0, t24 and t48 representthe ear thickness before and 24 or 48 hours after challengerespectively, in individual tests (T). The results were expressed as themean ± S.E.M. The level of significance between means of the groups wasobtained by Student's two-tailed t-test. Tables 1 and 2 show the resultsfrom 24 and 48 hours measurements respectively, from a representativeexperiment. The immunostimulating potency of the compound is reflectedin a significant difference in the increase in ear thickness as comparedto the control. Thus, in treated animals, after 24 h the response wasabout twice that of the control animals (15 μm compared to 8 μm, Table1).

                  TABLE 1                                                         ______________________________________                                        Ear thickness 24 hours after challenge of animals treated with the            indicated doses of di-L-lysinium-N,N'-diacetyl-L-cystinate                    or vehicle (NaCl).                                                            Dose            Diff.                                                         μmol/kg                                                                              N     T.sub.t24- T.sub.t0                                                                       S.E.M.                                                                              Sign.                                       ______________________________________                                        NaCl      10    8.25        0.56                                              0.03      10    15.00       0.42  ***                                         3.0       10    15.80       0.77  ***                                         ______________________________________                                         ***P < 0.001.                                                            

                  TABLE 2                                                         ______________________________________                                        Ear thickness 48 hours after challenge of animals treated with the            indicated doses of di-L-lysinium-N,N'-diacetyl-L-cystinate or                 vehicle (NaCl).                                                               Dose            Diff.                                                         μmol/kg                                                                              N     T.sub.t24- T.sub.t0                                                                       S.E.M.                                                                              Sign.                                       ______________________________________                                        NaCl      10    8.83        0.31                                              0.03      10    12.55       0.41  ***                                         3.0       10    13.20       0.28  ***                                         ______________________________________                                         ***P < 0.001.                                                            

The ability of the same test compound (la, R=lysine) to prolong orincrease the survival of rats suffering from tumours is illustrated bytwo different experiments with tumour inoculation in rats.

Two groups each consisting of ten rats of the Wistar Furth strain wereinoculated subcutaneously with 10⁴ mammary carcinoma cells per rat andtumour size was measured twice a week. In one group the rats weredrinking ordinary water and in the other group the test compounddissolved in water so as to result in a dose of 0,03 μmol/kg/day ofanimal weight. After 38 days one animal from the water-drinking groupwas still alive while from the group which was given the compound sevenanimals were alive.

In another experiment three groups each consisting of ten rats of theLewis strain were inoculated subcutaneously with 10⁶ glioma cells perrat and tumour size was measured twice a week. In one group the ratswere drinking ordinary water and in the second group the test compounddissolved in water so as to give a dose of 0,03 μmol/kg/day of animalweight, while in the third group the rats were drinking water containingLevamisole, a well-known cancer therapeutic drug also at a dose of 0,03μmol/kg/day of animal weight. After 119 days there was no surviving ratin the water-drinking group. In the group, which was given the testcompound, six rats were still alive while in the group, which receivedLevamisole, three rats were alive after 119 days.

In both experiments with tumours there was a lower frequency of tumourpositive animals and in both experiments there was an inhibition oftumour growth of the developed tumours in the groups receiving thecompound solubilized in water.

In another experiment two groups each consisting of eight SCID mice(mice with the immune defect Severe Combined Immunodeficiency Disease)were inoculated with 2*10³ mammary carcinoma cells and outgrowth wasmeasured twice a week. In one group the mice were drinking ordinarywater and in the second group the test compound dissolved in water so asto give a dose of 0.03 μmol/kg/day of animal weight. There were nosignificant differences between the groups concerning growth rate andincidence. This indicates that the compound has no direct effect on thetumour cells but acts via the immune apparatus.

In an animal model of lupus, experimental murine systemic lupuserythematosus (SLE), MRL Ipr/Ipr mice spontaneously develop lymphoidhyperplasia, dermatitis, arthritis and glomerulonephritis. The effect ofdi-L-lysinium-N,N'(-diacetyl-L-cystinate) on glomerulonephritis asproteinuria and hematuria and also the survival rate of the animals, hasbeen studied in this murine lupus model.Di-L-lysinium-N,N'-diacetyl-L-cystinate was administrated in thedrinking water and the mean dosage was calculated to 0.03 μmol/kg/day.The control mice received tap water. Both groups had free access to thedrinking water. The treatment started when the animals reached 8 weeksof age and continued until death or 46 weeks of age, when the study wasended.

The assessment of proteinuria and haematuria was performed by the usageof reagent strips, Eour-Test*, Boehringer Mannheim.

Upon administration of di-L-lysinium-N,N'-diacetyl-L-cystinate thesurvival rate was significantly improved compared to the control mice.The mortality rate for the untreated group (21 animals) reached 50%around 25 weeks of age. For the di-L-lysinium-N,N'-diacetyl-L-cystinatetreated MRL-Ipr/Ipr mice (12 animals) this mortality rate was notreached until week 44.

This form of treatment also significantly improved the score for bothproteinuria and haematuria measured as arbitrary units of the reagentstrips, compared to the untreated group.

These results show the immunomodulating potency of the compounds of thisinvention, particularly with regard to SLE.

WORKING EXAMPLES Pharmaceutical Formulations

The following examples are representative of pharmaceutical formulationsintended for different modes of local and systemic administration.

EXAMPLE 1 Pressurised Aerosol for Inhalation

The aerosol system is arranged so that each metered dose contains0.1-1.0 mg.

    ______________________________________                                        Compound Ia or Ib, micronized                                                                      1.0% w/w                                                 Sorbitan trioleate   0.7% w/w                                                 Trichloromonofluoromethane                                                                        24.4% w/w                                                 Dichlorotetrafluoroethane                                                                         24.4% w/w                                                 Dichlorodifluoromethane                                                                           49.5% w/w                                                 ______________________________________                                    

EXAMPLE 2 Powder Aersol for Inhalation Of Pure Substance

Pure substance prepared for inhalation from Turbuhaler. Each single dosecontains 0.1-1.0 mg.

    ______________________________________                                        Compound Ia or Ib, processed                                                                     0.1-1.0 mg                                                 ______________________________________                                    

EXAMPLE 3 Powder Aerosol for Inhalation

Each single dose contains 0.1-1.0 mg in a capsule.

    ______________________________________                                        Compound I, micronized                                                                              0.1-1.0 mg                                              Lactose               50      mg                                              ______________________________________                                    

EXAMPLE 4 Solution for Nebulising

The solution contains 1.0-10.0 mg/mL and 1-3 mL may be administered in asingle dose.

    ______________________________________                                        Compound             1.0-10.0 mg                                              Water for injection  to 1.0   mL                                              ______________________________________                                    

EXAMPLE 5 Tablets

Each tablet contains:

    ______________________________________                                        Compound I             0.1-100 mg                                             Maize starch           50      mg                                             Lactose                150     mg                                             Polyvidone             7       mg                                             Microcrystalline cellulose                                                                           20      mg                                             Magnesium stearate     2       mg                                             ______________________________________                                    

EXAMPLE 6 Oral solution

A single dose of 10 mL contains 10-100 mg.

    ______________________________________                                        Compound I            1-10   mg                                               Sorbitol 70%          150    mg                                               Glycerol              100    mg                                               Sodium benzoate       1      mg                                               Flavour               q.s.                                                    Water purified        to 1.0 mL                                               ______________________________________                                    

EXAMPLE 7 Tablet for Controlled Release

1 tablet:

    ______________________________________                                        Compound L            1-100   mg                                              Paraffin Special      145     mg                                              Lactose Powder        50      mg                                              Colloidal Silicon Dioxide                                                                           5       mg                                              Ethylcellulose 10 cps 13      mg                                              Ethanol 99,5 vol %    85      mg                                              Magnesium Stearate    2,5     mg                                              ______________________________________                                    

EXAMPLE 8 Granulate for Controlled Release

1 g of granulate:

    ______________________________________                                        Compound 1            1-100   mg                                              Ethylcellulose Dispersion                                                                           10      mg                                              Acetyltributylcitrate 0,5     mg                                              Eudragit L 100-55     55      mg                                              Triethylcitrate       5       mg                                              Talc                  30      mg                                              Water newly distilied 350     mg                                              Pellets, neutral      to 1000 mg                                              ______________________________________                                    

EXAMPLE 9 Solution for injection

1 mL in a single dose contains 1.0-10.0 mg

    ______________________________________                                        Compound Ia or Ib    1.0-10.0 mg                                              Sodium chloride      8.9-7.7  mg                                              Water for injection  to 1.0   mL                                              ______________________________________                                    

EXAMPLE 10 Cream for Topical Application

1 g of cream contains:

    ______________________________________                                        Compound I            0.1-1  mg                                               White soft paraffin   75     mg                                               Liquid paraffin       10     mg                                               Cetostearyl alcohol   75     mg                                               Cetomacrogol 1000     20     mg                                               Metagin               0.8    mg                                               Propagin              0.2    mg                                               Water, purified       to 1.0 g                                                ______________________________________                                    

EXAMPLE 11 Ointment for Topical Application

1 g of ointment contains:

    ______________________________________                                        Compound I            0.1-1  mg                                               Liquid paraffin       150    mg                                               White soft paraffin   to 1.0 g                                                ______________________________________                                    

EXAMPLE 12 Ophthalmic Solution

One dose of 2 drops contains 0.01-0.1 mg of compound I

    ______________________________________                                        Compound I             0.1-1  mg                                              Benzalkonium chloride  0.1    mg                                              Sodium chloride        9.0    mg                                              Water, sterile         to 1.0 mL                                              ______________________________________                                    

CHEMISTRY EXAMPLE 13 Di-L-lysinium-N,N'-diacetyl-L-cystinate (Ia; R═H₂N(COOH)CH(CH₂)₄ NH₃)

N-Acetyl-L-cysteine (22 mol, 3.59 kg) was dissolved in 2.6 L ofdeionized water. Sodium hydroxide 45% in water (22 mol, 1.92 kg) wasadded with stirring and temperature kept below 20° C. After adjustingthe temperature to 5° C., hydrogen peroxide (11.0 mol, 0.95 L) wascarefully added dropwise during 90 min. The temperature was not allowedto exceed 10° C. during this addition. To the resulting solution wasadded 9 L of activated strongly acidic cation exchanger. After stirringfor 10 minutes the pH was 2.0 and the ion exchanger was filtered off.The filtrate contained 9.65 mol of N,N'-diacetyl-L-cystine, asdetermined by HPLC using a standard prepared from the pure substance. Tothis crude solution was added L-lysine (19.3 mol, 3.17 kg). The thicksolution formed, was slowly added to 50 L of refluxing ethanolcontaining 0.23 kg of crystallinedi-L-lysinium-N,N'-diacetyl-L-cystinate. After the addition, the slurrywas allowed to cool and the crystals were filtered off. Washing withethanol (8 L) and drying (vacuum, 40° C.) for 12 hours afforded 5.36 kg(90%) of the title substance as a white crystalline solid.

Physical data: Mp: 210° C. (dec); α!_(D) ²⁵ =-70° (c=0.54, H₂ O); ¹H-NMR (D₂ O)δ:1.36-1.60 4H, m, Lys γCH₂ !, 1.73 4H, p, Lys δCH₂ !,1.84-1.96 4H, m, Lys βCH₂ !, 2.05 6H, s, CH₃ !, 2.95 2H, dd, CH₂ S!,3.02 4H, t, Lys εCH₂ !, 3.25 2H, dd, CH₂ S!, 3.76 2H, t, Lys αCH!, 4.502H, dd, CHN!¹³ C-NMR (D₂ Oδ:24.27; 24.80; 29.24; 32.72; 41.89; 42.78;52.96; 57.30; 176.53; 177.41; 179.74; Analysis: Calcd for C₂₂ H₄₄ O₁₀ N₆S₂, C:42.8 H:7.2 N:13.6 Found, C:42.6 H:7.4N:13.7; MS m/z=325 (MH⁺),m/z=471 (MLysH⁺), m/z=617 (MLys₂ H⁺).

EXAMPLE 14 Ethylenediaminium-N,N'-diacetyl-L-cystinate (Ib; R═H₃ NCH₂CH₂ NH₃)

To N,N'-Diacetyl-L-cystine (30.9 mmol, 10 g) dissolved in 20 mL of waterwas added ethylenediamine (61.8 mmol, 3.73 g) and ethanol (30 mL). Thesolution was concentrated to a thick paste which was redissolved in 80mL of ethanol. Crystallisation occurred after 2 hours stirring at 10°C.Filtration and drying gave 6.2 g (45%) of the title compound.

Physical data: Mp 185.2°-192.4° C. ¹ H-NMR (D₂ O)δ; 2.06 6H, s, CH₃ !,2.96 2H, dd, CH₂ S!, 3.26 2H, dd, CH₂ S!, 3.28 4H, s, H₃ N⁺ CH₂ CH₂ N⁺H₃ !, 4.50 2H, dd, CHN!.

¹³ C-NMR (D₂ O) δ: 24.78; 39.99; 42.74; 56.98; 176.55; 179.82; Analysis:Calcd C: 37.5, H:6.3, N:14.6, S:16.7. Found, C:37.3, H:6.8, N:15.3,S:15.2; MS m/z=385 M(H₂ NCH₂ CH₂ NH₂)H⁺ !

EXAMPLE 15 N,N'-dibenzylethylenediaminium-N,N'-diacetyl-L-cystinate (Ib;R═PhCH₂ NH₂ CH₂ CH₂ NH₂ CH₂ Ph)

To a 63% solution of N,N'-diacetyl-L-cystine in water (67 mmole, 21.8 g)was added N,N'-dibenzylethylenediamine (67 mmole, 16.0 g). Theexothermic reaction gave a slightly oily product which could berecrystallised from water to give 12.0 g (32%) of the title substance aswhite crystalline needles.

Physical data: Mp 163.8°-165.3° C. ¹ H-NMR (D₂ O) δ:2.04 6H,s,CH₃, 2.932H,dd,CH₂ S!, 3.22 2H,dd,CH₂ S!, 3.44 (4H,s,CH₂ NBn!, 4.27 4H,s,PhCH₂N!, 4.47 2H,dd,CHN!, 7.44-7.54 10H,m,Ph!. ¹³ C-NMR (D₂ O) δ:24.82,42.82, 45.71, 54.47, 56.99, 132.22, 132.58, 132.67, 133.52, 176.56,179.80. Analysis (monohydrate): Calcd, C: 53.6, H:6.6, N:9.6, S:11.0.Found, C:54.5, H:6.6, N:9.6, S:11.2.

EXAMPLE 16 Di-(1-adamantanaminium)-N,N'-diacetyl-L-cystinate (Ia; R═C(1) CH(3,5,7) CH₂ (2,4,6,8,9,10) NH₃ !

To N,N'-diacetyl-L-cystine (5.35 mmole, 1.73 g) dissolved in 5 mL ofwater was added 1-adamantanamine (10.7 mmole, 1.61 g). To the solutionwas dropwise added 60 mL of acetone. The resulting crystalline salt wasfiltered and dried in vacuo, yielding 2.3 g (67%) of the title compound.

Physical data: Mp 162° C., ¹ H-NMR (D₂ O)δ: 1.71 12H, broad dd, CH₂(4,6,10)!, 1.87 12H,d,CH₂ (2,8,9)!, 2.05 6H,s,CH₃ !, 2.17 6H, broad s,CH(3,5,7)!, 2.96 2H,dd,CH₂ S!, 3.26 2H,dd,CH₂ S!, 4.50 2H,dd,CHN!

EXAMPLE 17 Di-(N-benzyl-2-phenylethylaminium)-N,N'-L-cystinate (Ia;R═PhCH₂ CH₂ NH₂ CH₂ Ph)

To N,N'-diacetyl-L-cystine (28.7 mmole, 9.3 g) dissolved in 20 mL ofwater was added N-benzyl-2-phenylethylamine (57.4 mmole, 12.1 g). Thesolution was concentrated to a thick paste, from which the salt slowlycrystallized. The crystalline title compound was isolated and dried.

Physical data: Mp 87° C. ¹ H-NMR (D₂ O) δ:2.05 6H,s,CH₃ !, 2.952H,dd,CH₂ S!, 3.04 4H,t,PhCH₂ C!, 3.24 2H,dd,CH₂ S!, 3.33 4H,t,CH₂ NBn!,4.25 4H,s,PhCH₂ N!, 4.49 2H,dd,CHN!, 7.30-7.52 20H,m,Ph!

EXAMPLE 18 Piperazinium-N,N'-diacetyl-L-cystinate Ib; R═NH₂ (1,4) CH₂(2,3,5,6)!

To N,N'-diacetyl-L-cystine (4.60 mmole, 1.49 g) dissolved in 5 mL ofwater was added piperazine (4.60 mmole, 0.90 g). To the solution wasadded enough isopropanole to cause formation of an oil, which slowlysolidifies. The salt was isolated and dried.

Physical data: Mp>170° C. (dec.) ¹ H-NMR (D₂ O) δ:2.05 6H,s,CH_(3!),2.96 2H,dd,CH₂ S!, 3.26 2H,dd,CH₂ S!, 3.42 8H,s,CH₂ (2,3,5,6)!, 4.492H,dd,CHN!

EXAMPLE 19 Di-L-lysinium-N,N'-diacetyl-L-cystinate (Ia; R═H₂N(COOH)CH(CH₂)₄ NH₃)

N-acetyl-L-cysteine (37 mmol, 6.0 g) and L-lysine (37 mmol, 5.4 g) weredissolved in 10 mL of deionized water. Hydrogen peroxide (18 mmol, 1.5mL) was added dropwise while stirring, and the temperature was keptbelow 25° C. The solution was stirred for an additional 4 h. The viscoussolution was slowly added to 150 mL of refluxing ethanol containing 0.50g of crystalline di-L-lysinium-N,N'-diacetyl-L-cystinate. After additionthe solution was allowed to cool, and the crystals were filtered off.Washing with ethanol (20 mL), and drying (vacuum, 45° C.) for 24 hafforded 10.0 g (84%) of the title substance as a white crystallinesolid.

Physical data: Mp: 208° C.; α!²⁵ _(D) =-73°(c=0.54, H₂ O)

We claim:
 1. A method for the treatment of diseases which are due todefects in the immune system in mammals, which comprises administeringto a mammal in need of such treatment an effective amount of a salt ofan organic base and N,N'-diacetylcystine having the formula ##STR2## ora hydrate or a solvate thereof wherein the organic base, R³⁰ or R²⁺, isselected from the group consisting of lysinium, ethylenediaminiumN,N'-dibenzylethylenediaminium, N-benzy2-phenylethylaminium and1-adamantanaminium and wherein the disease is selected from the groupconsisting of chronic bronchitis, rheumatoid arthritis, hepatitis,asthma, rhinitis, atherosclerosis, HIV infection and AIDS.
 2. The methodaccording to claim 1 wherein the effective amount of the salt isadministered in solid form as a dried powder from a pressurized meterdose inhaler.
 3. The method according to claim 1 wherein the salt iscrystallized.